Project summary: Microglia are immune cells that migrate to the brain, taking up residency early in development, and constitute the first line of defense against brain infection, disease and injury. However, recent studies have shown that microglia are essential participants in the physiological brain and play critical roles in normal functions such as network development, synaptic plasticity and adult neurogenesis. While the understanding of the dual roles of microglia in health and disease has stimulated numerous lines of research, the majority of studies investigating microglial dynamics during both pathological and physiological events have been carried out in anesthetized animals, which exhibit a slow-wave sleep-like state. Given that slow-wave sleep is a critical component in microglia-implicated processes such as facilitating synaptic development, network maintenance, and post-injury recovery, this leaves open the possibility that microglia have different roles in the brain depending on arousal state. Here we will investigate the hypothesis that noradrenergic signaling during wakefulness dampens microglial dynamics. Understanding how arousal state modulates microglial behavior will be critical to understanding how these cells contribute to normal brain function and disease progression. It is well established that sleep helps maintain brain homeostasis and facilitates normal neurological processes. Disruptions in sleep quality can impair behavioral performance, inhibit learning and memory, and lead to negative outcomes in numerous neuropathologies. While the biological basis for sleep?s effects on the brain is currently unknown, we posit that homeostatic and immune functions of microglia during sleep states play an important role. Microglia are highly motile under basal physiological conditions, dynamically surveying synapses and contributing to changes in synaptic strength during both physiological (e.g. learning and memory) and pathological (e.g. excitotoxicity) events. Our preliminary experiments show that microglial motility is significantly reduced in awake animals, suggesting that microglial surveillance occurs primarily during sleep. Here we will explore this further by comparing the dynamics of microglia and their ability to interact with synaptic structure in vivo in anesthetized, awake and naturally sleeping animals in Aim1. In Aim 2 we will determine whether noradrenergic signaling is responsible for the effects of wakefulness on microglial dynamics and whether noradrenaline signals directly to microglia or through changes in extracellular space. The two aims are designed to provide a thorough investigation into the role arousal plays on microglial dynamics.